French Suite No. 5 by J. S. Bach , with or without inaudible high-frequency components, while performing a visual vigilance task. High-alpha (10.5–13 Hz) and low-beta (13–20 Hz) EEG powers were larger for the excerpt with high-frequency components than for the excerpt without them. Reaction times and error rates did not change during the task and were not different between the excerpts. The amplitude of the P3 component elicited by target stimuli in the vigilance task increased in the second half of the listening period for the excerpt with high-frequency components, whereas no such P3 amplitude change was observed for the other excerpt without them. The participants did not distinguish between these excerpts in terms of sound quality. Only a subjective rating of inactive pleasantness after listening was higher for the excerpt with high-frequency components than for the other excerpt. The present study shows that high-resolution audio that retains high-frequency components has an advantage over similar and indistinguishable digital sound sources in which such components are artificially cut off, suggesting that high-resolution audio with inaudible high-frequency components induces a relaxed attentional state without conscious awareness.High-resolution audio has a higher sampling frequency and a greater bit depth than conventional low-resolution audio such as compact disks. The higher sampling frequency enables inaudible sound components (above 20 kHz) that are cut off in low-resolution audio to be reproduced. Previous studies of high-resolution audio have mainly focused on the effect of such high-frequency components. It is known that alpha-band power in a human electroencephalogram (EEG) is larger when the inaudible high-frequency components are present than when they are absent. Traditionally, alpha-band EEG activity has been associated with arousal level. However, no previous studies have explored whether sound sources with high-frequency components affect the arousal level of listeners. The present study examined this possibility by having 22 participants listen to two types of a 400-s musical excerpt of  High-resolution audio has recently emerged in the digital music market due to recent advances in information and communications technologies. Because of a higher sampling frequency and a greater bit depth than conventional low-resolution audio such as compact disks (CDs), it provides a closer replication of the real analog sound waves. Sampling frequency means the number of samples per second taken from a sound source through analog-to-digital conversion. Bit depth is the number of possible values in each sample and expressed as a power of two. A higher sampling frequency makes the digitization of sound more accurate in the time-frequency domain, whereas a greater bit depth increases the resolution of the sound. What kind of advantage does the latest digital audio have for human beings? This question has not been sufficiently discussed. The present investigation used physiological, behavioral, and subjective measures to provide evidence that high-resolution audio affects human psychophysiological state without conscious awareness.The higher sampling frequency enables higher frequency sound components to be reproduced, because one-half of the sampling frequency defines the upper limit of reproducible frequencies (as dictated by the Nyquist–Shannon sampling theorem). However, in conventional digital audio, sampling frequency is usually restrained so that sounds above 20 kHz are cut off in order to reduce file sizes for convenience. This reduction is based on the knowledge that sounds above 20 kHz do not influence sound quality ratings  and do nIn contrast to this conventional digital recording process in which inaudible high-frequency components are cut off, high-resolution music that retains such components has been repeatedly shown to affect human electroencephalographic (EEG) activity . This efPrevious studies have examined the effect of inaudible high-frequency components on EEG activity while listening to music under resting conditions. It has been shown that EEG alpha-band (8–13 Hz) frequency power is greater for high-resolution music with high-frequency components than for the same sound sources without them . The effA study using positron emission tomography (PET) revealed that the brainstem and thalamus areas were more activated when hearing full-range as compared with high-cut sounds . BecauseAnother line of research suggests a link between cognitive function and alpha-band as well as beta-band EEG activities. Alpha-band EEG activity is thought to be associated not only with arousal and vigilance levels  but alsoBeta-band power is broadly thought to be associated with motor function when it is derived from motor areas . MoreoveWhat kind of advantage does high-resolution audio with inaudible high-frequency components have for human beings? What remains unclear is what kind of psychophysiological states high-resolution audio induces, along with the corresponding increase in alpha- and beta-band EEG activities. To monitor listeners’ arousal level, we asked participants to listen to a musical piece while performing a visual vigilance task that required sustained attention in order to continuously respond to specific stimuli. Two types of high-resolution audio of the same musical piece were presented using a double-blind method: With or without inaudible high-frequency components.EEG was recorded along with other psychophysiological measures: Heart rate (HR), heart rate variability (HRV), and facial electromyograms (EMGs). The former two measures index autonomic nervous system activities. HRV contains two components with different frequency bands: High frequency , and low frequency . HF and LF activities are mediated by vagal and vagosympathetic activations, respectively . The LF/In the present study, physiological, behavioral, and subjective measures were recorded to examine what kind of advantage high-resolution audio with inaudible high-frequency components has. Specifically, we were interested in how the increase in alpha- and beta-band EEG activities is associated with listeners’ arousal and vigilance level. Using a double-blind method, two types of high-resolution audio of the same musical piece (with or without inaudible high-frequency components) were presented while participants performed a vigilance task in the visual modality.M = 20.6 years) did not report any known neurological dysfunction or hearing deficit. They were right-handed according to the Edinburgh Inventory (M = 84.1 ± 12.8). All reported to have correct or corrected-to-normal vision. Eight participants had the experience of learning musical instruments for a few years, but none of them were professional musicians. The Research Ethics Committee of the Graduate School of Integrated Arts and Sciences in Hiroshima University approved the experimental protocol.Twenty-six student volunteers at Hiroshima University gave their informed consent and participated in the study. Four participants had to be excluded due to technical problems. The remaining 22 participants  was selected. In the present study, this portion was played twice to produce a 400-s excerpt. The original (full-range) excerpt is rich in high-frequency components. A high-cut version of the excerpt was produced by removing such components using a low-pass finite impulse response digital filter with a very steep slope . This linear-phase filter does not cause any phase distortion. Although the filter produces very small ripples (1.04∗10-2 dB), they are negligible and it is unlikely to affect auditory perception. Sounds were amplified using AI-501DA  controlled by dedicated software on a laptop PC. Two loudspeakers with high-frequency tweeters  were located 1.5 m diagonally forward from the listening position. The sound pressure level was set at approximately 70 dB (A). Calibration measurements at the listening position ensured that the full-range excerpt contained abundant high-frequency components and that the high-frequency power of the high-cut excerpt  did not differ from that of background noise. The average power spectra of the excerpts are available at http://links.lww.com/WNR/A279 as Supplemental Digital Content of The present study used the same materials that were used in An equiprobable visual Go/NoGo task was conducted using a cathode ray tube (CRT) computer monitor (refresh rate = 100 Hz) in front of participants. A block consisted of 120 visual stimuli: 60 targets  and 60 non-targets (‘O’) in a randomized order. The visual stimuli were 200 ms in duration and presented with a mean stimulus onset asynchrony (SOA) of 5 s (range = 3-7 s). Button-press responses with the left and right index fingers were required to ‘T’ and ‘V’ (or ‘V’ and ‘T’) respectively, as quickly and accurately as possible.The study was conducted using a double-blind method. Participants listened to two versions of the 400-s musical excerpt (with or without high-frequency components) while performing the Go/NoGo task. Participants also performed the task under silent conditions for 100 s before and after music presentation (pre- and post-music periods). The presentation order of the two excerpts was counterbalanced across the participants. EEG, HR, and facial EMGs were recorded during task performance. After listening to each excerpt, participants completed a sound quality questionnaire consisting of 10 pairs of adjectives and then reported their mood states on the Affect Grid  and multPsychophysiological measures were recorded with a sampling rate of 1000 Hz using QuickAmp . Filter bandpass was DC to 200 Hz. EEG was recorded from 34 scalp electrodes  according to the extended 10–20 system. Four additional electrodes  were used to monitor eye movements and blinks. EEG data were recorded using the average reference online and re-referenced to the digitally linked earlobes (A1–A2) offline. EEG data were resampled at 250 Hz and were filtered offline . Ocular artifacts were corrected using a semi-automatic independent component analysis method implemented on Brain Vision Analyzer 2.04 (Brain Products). The components that were easily identifiable as artifacts related to blinks and eye movements were removed.Heart rate was measured by recording electrocardiograms from the left ankle and the right hand. The R–R intervals were calculated and converted into HR in bpm. For facial EMGs, electrical activities over the zygomaticus major and corrugator supercilii regions were recorded using bipolar electrodes affixed above the left brow and on the left cheek, respectively . The EMG2) of the following frequency bands were calculated: Delta (1–4 Hz), theta (4–8 Hz), low-alpha (8–10.5 Hz), high-alpha (10.5–13 Hz), low-beta (13–20 Hz), high-beta (20–30 Hz), and gamma (36–44 Hz). The square root of the total power (μV) was used for statistical analysis, following the procedure of previous studies  was divided into six 100-s epochs. For EEG analysis, each 100-s epoch was divided into 97 2.048-s segments with 1.024 s overlap. Power spectrum was calculated by Fast Fourier Transform with a Hanning window. The total powers (μV studies . The sca2) were calculated for LF (0.04–0.15 Hz) and HF (0.15–0.4 Hz) bands, and LF/HF power ratio was obtained. The square roots of LF and HF (ms) were used for statistical analysis.Heart rate variability analysis was done by using Kubios HRV 2.2 . The lasFor ERP analysis, the total 400-s listening period was divided into two 200-s epochs, to secure a reasonable number of Go trials (around 20). Silent periods (pre- and post-music epoch) were not included in the calculation. Those trials found to have Go omissions, Go misses (incorrect hand response to ‘T’ or ‘V’), or NoGo responses (commission errors) were excluded from further processing steps. Go and NoGo responses were separately averaged to produce ERPs. Epochs (200 ms before stimulus presentation until 1000 ms after the presentation) were baseline corrected (-200 ms until 0 ms). The peak of a P3 wave was identified within a latency range of 350-500 ms at Pz where P3 amplitude is dominant topographically.post hoc multiple comparisons of means, the comparison-wise level of significance was determined by the Bonferroni method.Each measure was subjected to a repeated measures analysis of variance (ANOVA) with sound type (full-range vs. high-cut) and epoch  as factors. To compensate for possible type I error inflation by the violation of sphericity, multivariate ANOVA solutions are reported . The sigFigure 1 shows the EEG amplitude spectrogram for the four regions in the silent conditions (pre- and post-music periods). Although participants were performing a visual vigilance task with eyes opened, a peak around 10 Hz appears clearly. The amplitude of the peak appears to be increased after listening to music, in particular after listening to the full-range version of the musical piece.Figure 2 shows the time course and scalp topography of high-alpha EEG (10.5–13 Hz) and low-beta EEG (13–20 Hz) bands. For EEG measures, a Sound Type × Epoch × Anterior-Posterior × Hemisphere ANOVA was conducted for each frequency band. Significant effects of sound type were found for both bands. For other frequency bands, only the theta EEG band (4-8 Hz) power showed a significant Sound Type × Anterior-Posterior × Hemisphere interaction, F = 5.37, p = 0.031, F = 7.06, p = 0.001, F = 12.63, p = 0.002, F = 6.99, p = 0.015, Post hoc tests revealed that high-alpha EEG power was greater for the full-range excerpt than for the high-cut excerpt and that the sound type effect was found for the left but not right hemisphere at the 200-300-s epoch. No effects of sound type were obtained at the epochs before 200 s. The main effect of anterior-posterior was also significant, F = 15.67, p = 0.001, For high-alpha EEG band, the Sound Type × Epoch × Hemisphere interaction was significant, F = 4.49, p = 0.046, F = 5.43, p = 0.030, F = 7.07, p = 0.015, F = 5.26, p = 0.032, F = 5.27, p = 0.032, F = 3.94, p = 0.060, Figure 2 shows that the difference between the full-range and high-cut excerpts seems to be more prominent at later epochs. Two-tailed t-tests revealed significant differences between the two excerpts at the 200-300-s, 300-400-s, and post epochs, ts(21) > 2.37, ps < 0.027; p > 0.114 at the epochs before 200 s.For low-beta EEG band, the Sound Type × Anterior-Posterior × Hemisphere interaction and the main effect of sound type effect were significant, Figure 3 shows grand mean ERP waveforms and the scalp topography of the Go and NoGo P3 amplitudes. The mean number of averaged trials was 18.6 (range = 13-20). Although this is less than an optimal number of averages for P3  = 4.39, p = 0.049, F = 2.64, p = 0.119, Post hoc tests revealed that Go P3 amplitude increased from the 0-200 s to the 200-400 s epoch for the full-range excerpt, whereas Go P3 amplitude did not change for the high-cut excerpt. The main effect of epoch was significant for the NoGo condition, F = 13.39, p = 0.001, For the P3 amplitude, a Sound Type × Epoch ANOVA was conducted for Go and NoGo stimulus conditions separately. A significant interaction was found for the Go condition, F = 5.01, p = 0.036, Similar ANOVAs were conducted for latencies. No significant main or interaction effects of sound type were found. The main effect of epoch was significant for the Go stimulus condition, One of the reviewers questioned about the effects of sound type on the Nogo N2 . We condM = 98.6%, 95.8-100%; full-range: M = 97.9%, 95.0-99.2%). Figure 4 shows the time course of mean Go reaction times, HR, and facial EMGs , and the HRV components for the last 300-s epoch of the musical excerpts. For the corrugator supercilii, a Sound Type × Epoch ANOVA showed a significant main effect of epoch, F = 5.69, p = 0.003, Participants performed the vigilance task with considerable accuracy  = 3.13, p = 0.005. Participants provided higher inactive pleasantness scores under the full-range than under the high-cut excerpt. Figure 5 shows the mean sound quality ratings for the full-range and high-cut musical excerpts. No significant differences were found between the two types of audio source for any adjective pairs, ts(21) < 1.92, ps > 0.069. The correct rate of the forced choices was 41.0%, which did not exceed chance level .High-resolution audio with inaudible high-frequency components is a closer replication of real sounds than similar and indistinguishable sounds in which these components are artificially cut off. It remains unclear what kind of advantages high-resolution audio might have for human beings. Previous research in which participants listened to high-resolution music under resting conditions have shown that alpha and low-beta EEG powers were larger for an excerpt with high-frequency components as compared with an excerpt without them . The preWe found greater high-alpha (10.5–13 Hz) and low-beta (13–20 Hz) EEG powers for the excerpt with high-frequency components as compared with the excerpt without them. The effect appeared in the latter half of the listening period (200-400 s) and during the 100-s period after music presentation (post-music epoch). Furthermore, for full-range sounds compared with high-cut sounds, Go trial P3 amplitude increased, and subjective relaxation scores were greater. Because task performance did not change across musical excerpts, with no difference in self-reported arousal, the effects of high-resolution audio with inaudible high-frequency components on brain activities should not reflect a decrease of listeners’ arousal level. These findings show that listeners seem to experience a relaxed attentional state when listening to high-resolution audio with inaudible high-frequency components compared to similar sounds without these components.It has been shown that listening to musical pieces increases EEG powers of theta, alpha, and beta bands , and thaThe present study shows that not only high-alpha and low-beta EEG powers but also P3 amplitude increased in the last half of the listening period (200-400 s). Alpha-band EEG activity and P3 amplitude have been shown to be positively correlated, in such a way that prestimulus alpha directly modulates positive potential amplitude in an auditory equiprobable Go/NoGo task . P3 amplAs a mechanism underlying the effect of inaudible high-frequency sound components, we speculate that the brain may subconsciously recognize high-resolution audio that retains high-frequency components as being more natural, as compared with similar sounds in which such components are artificially removed. A link between alpha power and ratings of ‘naturalness’ of music has been reported. When listening to the same musical piece with different tempos, alpha-band EEG power increased for excerpts that were rated to be more natural, the ratings of which were not directly related to subjective arousal . As highOur findings have some limitations. First, because we used only a visual vigilance task, it is unclear whether high-resolution audio can improve performance on tasks that involve working memory and long-term memory. Because a vigilance task is relatively easy, our participants were able to sustain high performance. Other research using an n-back task requiring memory has shown that high-resolution audio also enhances task performance . Future Second, the underlying mechanism of how inaudible high-frequency components affect EEG activities cannot be revealed by the current data. It is noteworthy that presenting high-frequency components above 20 kHz alone did not produce any change in EEG activities . TherefoThird, it remains unclear why there was a time lag until the effects of high-resolution audio on brain activity show up, and why this effect was maintained for 100 s after music stopped. A possible reason is that, as mentioned above, sufficiently long exposure is needed for the effects of inaudible high-frequency components. Another possibility is that listening to music has psychophysiological impact through the engagement of various neurochemical systems . HumoralFourth, the present study did not manipulate the sampling frequency and the bit depth of digital audio. High-resolution audio is characterized not only by the capability of reproducing inaudible high-frequency components but also by more accurate sampling and quantization  as compared with low-resolution audio. If the naturalness derived by a closer replication of real sounds affects EEG activities, the sampling frequency and the bit depth would do too regardless of whether the real sounds feature high-frequency components. This idea would be worth examining in future research.In summary, high-resolution audio with inaudible high-frequency components has some advantages over similar and indistinguishable sounds in which these components are artificially cut off, such that the former type of digital audio induces a relaxed attentional state. Even without conscious awareness, a closer replication of real sounds in terms of frequency structure appears to bring out greater potential effects of music on human psychophysiological state and behavior.This study was carried out in accordance with the recommendations of The Research Ethics Committee of the Graduate School of Integrated Arts and Sciences in Hiroshima University. All participants gave written informed consent in accordance with the Declaration of Helsinki. The protocol was approved by The Research Ethics Committee of the Graduate School of Integrated Arts and Sciences in Hiroshima University.RK and HN planned the experiment, interpreted the data, and wrote the paper. RK collected and analyzed the data.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The aim of this study was to determine arch width changes during maxillary first premolars extraction and non-extraction treatment in patients with Class II division 1 malocclusion. Dental casts of 91 Class II division 1 patients  were evaluated. The minimum age of the subjects at the beginning of treatment was above 16 years. 48 patients were treated with extraction of the maxillary first premolars and 43 patients were treated without extraction. Pre- and post-treatment maxillary and mandibular inter-canine and inter-molar arch widths were measured.At the end of treatment, maxillary and mandibular inter-canine widths of both groups increased significantly. The maxillary inter-molar width decreased in the extraction group and increased in the non-extraction group. The mandibular inter-molar width increased significantly in both groups. No significant differences were observed between males and females.The results of this study indicated that there was a tendency for an increase in arch width during both the extraction and non-extraction treatment except maxillary inter-molar width in the extraction cases. Key words:Dental arch, malocclusion, angle Class II, tooth movement, extraction. One of the major issues of concern and debate in orthodontics is the stability of achieved result . HoweverAesthetics is one of the main goals of orthodontic treatment. Narrower dental arches in extraction treatment, when compared with non-extraction, have been criticized in previous studies . Howeveret al. .Based on 80% power and significance level of 5% , and con1. All cases were originally diagnosed as having mild to moderate skeletal Class II division 1 malocclusion.2. None of the cases had congenital anomalies, significant facial asymmetries, or congenitally missing teeth.3. All cases were above 16 years of age and all were in the permanent dentition.4. All cases received no palatal expansion, functional appliance, orthognathic surgery or fixed prosthodontic therapy5. All cases had overbite of 5% to 40% and mandibular arch crowding of ≤4 mm.6. All cases were treated with fixed preadjusted (0.022-inch bracket slot) technique with class II elastics for non-extraction and space closure with sliding for extraction cases.7. A clinically acceptable occlusion was established after active treatment i.e., a Class I canine relationship, an overbite between 10% and 25%, and well-aligned and inter-digitated arches.8. Plaster dental casts were taken before and after orthodontic treatment.These criteria were adopted to insure that post-treatment changes were not caused by poor treatment results.-Study casts analysis Four arch width measurements were recorded from each subject’s dental casts using a digital calliper and recording the data to the nearest 0.1 mm. These measurements included: (A) maxillary inter-canine width between the height of contour points on the main buccal ridge located at the cervical third of the canines, (B) maxillary inter-molar width between the height of contour points located gingival to buccal grooves of the first molars, (C) mandibular inter-molar width between the height of contour points located gingival to main buccal pits of the first molars and (D) mandibular inter-canine width between the height of contour points on the buccal ridge located at the cervical third of the canines . A paired sample t-test was used to evaluate the treatment changes within each group. To compare the changes in the extraction and non-extraction groups for both males and females, independent student t-test was used. Statistical significance level was established at P>0.05). The sex distribution in groups was not significantly different (P>0.05). Almost perfect intra-examiner reliability for all examiners was determined ; inter-examiner reliability was also perfect (ICC = 0.92).A total of 91 Class II division 1 patients  were included in the study. At the start of treatment, there was no statistically significant difference between arch widths of both groups . This was the same for the non-extraction group  (P=0.001)  . This inP=0.001) . CompariP=0.001) .P<0.001) (P>0.05). In the non-extraction group, inter-molar width significantly increased during the same period (P<0.001) (Inter-molar width changes were in the opposite direction comparing the extraction and non-extraction groups (-0.56 ±0.64 mm for the extraction group and 2.18 ±0.85 mm for the non-extraction group) (P<0.001) . In the P<0.001) . SimilarP<0.001) .-Evaluation of the lower archP>0.05)  .P<0.001) (P>0.05). Inter-molar width increased 1.39 ± 1.06 mm in the extraction group and 1.58 ± 0.93 mm in the non-extraction group (P>0.05) (Mandibular inter-canine and inter-molar widths significantly increased at the end of treatment (P<0.001) . The mea(P>0.05) . The tre(P>0.05) .-Sex comparison P>0.05). However, in general, dimensional increases or decreases tend to be greater in females.The differences between males and females were not statistically significant , an increase of about 2.5 mm in transpalatal width between the upper first molars has been reported . Other IThe results of this study indicated that there is a tendency for an increase in arch width during both the extraction and non-extraction treatment in Class II division 1 malocclusions except maxillary inter-molar width in the extraction cases. In general, when comparing the findings of this study with those of other studies, it seems that the direction of treatment changes is similar in arches treated with extraction in the Class II cases. This comparison is also true for arches treated with non-extraction method. Nevertheless, it should be borne in mind that when treating a patient with extraction in one arch and non-extraction in the other arch, the two arches can influence each other. One might argue that there might be selection bias at the start of the study because of its retrospective design. However, it should be mentioned that the records of all consecutive patients, from 2000 to 2014, fulfilling the inclusion criteria were evaluated in order to avoid selection bias. Further prospective studies controlling for probable confounding factors are warranted.In the extraction cases, if we assume that the arch dimensional changes are because of tooth movement into narrower or wider parts of the arch, then different anchorage values and different arch forms will cause different changes in arch width between identical teeth. This needs to be investigated in future studies.


P < 0.05). PI  score was the only independent prognostic predictor for OS and PFS of HCCs in the training set. These results were further confirmed in a validation set. In conclusion, differentially expressed serum miRNAs can be helpful for predicting survival in HCCs.To establish serum microRNA profiles as prognostic biomarkers in hepatocellular carcinoma patients (HCCs), we used deep sequencing to screen serum microRNAs in a discovery set. Twelve up-regulated serum miRNAs were selected for qPCR analysis in a training set. MiR-192-5p and miR-29a-3p were identified and associated with HCC prognosis. HCCs with high concentrations of miR-192-5p and miR-29a-3p had poorer overall survival (OS) and progression-free survival (PFS) than those with low concentrations. We calculated a prognostic index (PI) score and classified patients into low-, medium- and high-risk groups. OS and PFS among the 3 groups from the training set were significantly different (all  Hepatocellular carcinoma (HCC) is the most common and highly malignant hepatoma, the fifth most common cancer, and the third most common cause of cancer-related death in the world , 2. SuchThe discovery of microRNAs (miRNAs) has opened new avenues for cancer diagnosis, prognosis and prediction of treatment response . miRNA sIn fact, differential expression of several miRNAs in the serum, including miR-16, miR-122, miR-21, miR-223, miR-25, miR-375 and let-7f in patients with HCC, patients with hepatitis B, and healthy individuals has been reported recently , 19. HowHere, we describe the differential expression of serum miRNAs in 50 hepatitis B virus (HBV)-positive HCCs and 50 healthy controls (HCs) using deep sequencing technology in a discovery set. We used these results to establish a risk model in a training set of 74 HBV-positive HCCs and further validated the risk model in a validation set of 100 HBV-positive HCCs.To derive optimal serum miRNA profiles, we carried out high-throughput next-generation sequencing of 50 HCC samples and 50 HC samples in the discovery stage. All annotated small RNAs are explained in P values < 0.05), 12 up-regulated and 7 down-regulated miRNAs with significantly different expression levels between HCCs and HCs were identified  were differentially altered between 74 HCCs and 60 HCs in the training set (see in the P = 0.001) and PFS (P = 0.001) than those with low concentrations of miR-192-5p  and PFS (P = 0.023) than those with low concentrations of miR-29a-3p , tumor rize, vascular invasion, miR-192-5p and miR-29a-3p were correlated with the survival of HCCs see . SubsequP = 0.0001) and 0.71 , respectively  in the training set, as shown in Table y Figure −3B.The median levels of the 2 serum miRNAs were significantly higher in HCCs than in control groups  Figure −3D. The In HCC patients, a decrease of the 2 serum miRNAs concentration between pre- and post-operated HCCs was observed in HCCs serum in the training set Figure .OS score, while another model combining BCLC stage, miR-192-5p and AFP was analyzed to calculate the PIPFS score. The scores were calculated by the following equations  + 1.45 (0 or 1) + 1.81(0 or 1); PIPFS = 1.89 (0 or 1) + 1.07 (0 or 1)+ 0.69 (0 or 1). The PIOS and the PIPFS score had a range of 0−5.45 and 0−3.65, respectively. Consequently, patients with low-, medium- and high-risk had OS/PFS-predictor scores of < 1.81/0.69, 1.81−3.64/0.69−1.89 and > 3.64/1.89.Finally, 3 variables, including BCLC stage, miR-192-5p and miR-29a-3p, were analyzed in a multivariate risk model to calculate the PIP = 0.000; P = 0.000) and PFS  than those in the medium- and high-risk group, and HCCs in the medium-risk group had longer OS (P = 0.038) and PFS (P = 0.009) than those in the high-risk group   was the only significantly independent prognostic predictor for OS  and PFS  of HCCs in the training set.HCCs in the low-risk group had significantly longer OS  Figure −5D. MultP = 0.000; P = 0.000) and PFS  than those in the medium- and high-risk group, and HCCs in the medium-risk group had longer OS (P = 0.016) and PFS (P = 0.004) than those in the high-risk group   and PFS  of HCCs by multivariate analyses.To validate the prognostic value of the PI score, we quantified the levels of serum miR-192-5p and miR-29a-3p in a validation set of 100 HCCs and 70 HCs samples. We then calculated the risk score for each patient and classified patients into high-, medium- and low-risk groups using the same formulas and method as used in the training set. Similar to what was seen in the training set, HCCs in the low-risk group had significantly longer OS  Figure −6D, and In HCC patients, an increase in miR-192-5p and miR-29a-3p staining intensity was observed in HCC tissues compared with corresponding normal adjacent tissues by FISH Figure .We identified miR-192-5p and miR-29a-3p as inversely associated with survival and progression in HCCs. We found that a combination of miRNA expression levels and clinical variables, had a better prognostic value than BCLC stage alone.In the clinic, HCCs with the same stage who receive similar treatment regimens often show large variations in disease outcome, indicating that the current staging system may have reached its limitations for prognostic assessment. Staging in HCC patients is challenging because it requires staging and grading of both tumor and the chronic liver disease. Therefore, novel markers that reflect the aggressiveness of the disease would complement the current set of prognostic and treatment algorithms.Circulating miRNAs are stably detectable in serum or plasma , 23, 24 Several publications suggest significant roles for miRNAs in the evaluation of cancer prognosis and diagnosis, such as lung cancer and HCC –30. In tLogistic regression with penalized estimates may be used to develop prognostic models for binary outcomes, especially when limited data are available. Discrimination, calibration, and overall performance were taken into account in the construction of models . ROC curDeep sequencing can be focused on both discovering novel miRNAs and acquiring a highly quantitative estimate of known individual miRNA species . DevelopDecrease of both miR-192-5p and miR-29a-3p has been shown in HCC cells in several studies , 38–40, The American Association for the Study of Liver Diseases Practice Guidelines (July 2010) discarded AFP for surveillance and diagnosis. Therefore, there is a need for novel markers that would serve as a reliable method for early detection and postsurgical progression. This need might be well fulfilled with serum miR-192-5p and serum miR-29a-3p as alternative biomarkers which may be particularly helpful for AFP-negative HCCsCaenorhabditis elegans [However, there are some limitations in our study. First, in the discovery stage, if we had compared the expression of serum miRNAs of the controls with that of the prognosis well and the poor HCC patients, it would be better in experimental design. Second, currently no established internal reference exists for circulating miRNAs. Non-human spike-in miRNAs, such as from  elegans , 43 haveIn conclusion, distinct serum miRNA profiles associated with survival exist in HCCs. Additionally, these serum miRNAs may be a robust predictor of prognosis for HCC, which may lead to more personalized therapy. Moreover, deep sequencing followed by qPCR validation may be a successful strategy to determine biomarkers in circulation.All samples were obtained from confirmed cases of HCC at the time of diagnosis from the Sun Yat-sen University Cancer Center (SYSUCC) . For the discovery stage of this study, we screened serum miRNAs in a set of 50 HCCs and 50 HCs consecutively obtained between January 2009 and April 2009. For subsequent study, , we used randomly selected samples obtained between May 2009 and December 2013 from 174 HCCs, 43cirrhosis patients and 130 HCs to test and validate miRNA expression at SYSUCC. HCs were individuals who came in for a routine physical examination and were found to be cancer-free. The characteristics of the 174 HCC patients are summarized in Table ® LS reagent  following the manufacturer's recommendations. More detailed information is provided in the Venous blood was incubated at room temperature for 1 hr, centrifuged at 500 g for 10 min and then 10,000 g for 30 min at 4°C to separate the serum. The supernatant was transferred to fresh RNase-free tubes and stored at −80°C until use. We extracted total RNA using TRIzolWe created 2 small RNA libraries according to the protocol published by Morin et al. . To fullDetailed information is provided in the To identify miRNAs differentially expressed between HCCs and HCs in the discovery stage, we used the Bayesian method developed by Audic and Claverie . miRNAs ®Green based quantitative PCR (qPCR) reactions were performed on the 7500 Real-time PCR systems  to validate 12 mature up-regulated miRNAs screened by sequencing  protocol were optimized and performed. More details are provided in the To study the spatial and temporal expression of miRNAs with high sensitivity and resolution, the miRNA fluorescein To compare the sequencing data of the HCCs and HCs, we used the Wilcoxon–Mann–Whitney test. For the data obtained by qPCR, either the Mann-Whitney unpaired test or the Kruskal–Wallis test was used for the comparison between HCC and control . We compared the serum miRNAs concentration of pre- and post-operated HCCs using Wilcoxon matched-pairs signed rank test. The levels of all miRNAs with significance values < 0.05 were considered statistically significant.We defined overall survival (OS) and progression free survival (PFS) as the period between the date of diagnosis and recurrence/metastasis, death, or last day of follow-up. Survival curves were estimated by use of the Kaplan–Meier method with the log-rank test. We used a multivariate Cox regression analysis to investigate whether the selected miRNAs were independent prognostic factors of OS and PFS in HCCs.For each miRNA expression profile, we plotted the sensitivity and specificity of each outcome as a predictor of death for HCC with receiver operating characteristic (ROC) curve analysis. We used ROC curves as an accuracy index for assessing the predictive power as well as evaluating the diagnostic performance of each variable. We used the ROC curves to select cutoff scores for dichotomizing each predictor according to the maximum area under the ROC curve (AUC). The cutoff scores were generated with MedCalc software, version 12.2.1.Y = 1 | X} = β0 + Σixiβ [Y is a binary outcome variable (0 or 1), β0 is an intercept, and denotes the regression coefficients associated with the iβ design matrix X of covariables i. Specifically, in our study we calculated the PI score according to the formula: PI = β1x1 + β2x2 + β3x3, where β is a variable assignment indicated in univariate Cox regression analyses for HCCs in the training set . In general, we defined the PI score in terms of the logistic regression model: PI = logit{ + Σβixi , where Y set see . A ROC cP values of < 0.05 were considered to indicate statistical significance. Statistical analyses were carried out by use of SPSS 17.0.Two-sided


Lindera benzoin L. Blume (northern spicebush). Covering all seven phenoseasons of a deciduous forest, this work utilized a line quantum sensor to measure the variation in subcanopy light levels under all sky conditions at different times of the day. A total of 4,592 individual subcanopy measurements of photosynthetic photon flux density  were taken as 15-second spatially-integrated one-meter linear averages to better understand the dynamism of light exposure to L. benzoin. Both open  and subcanopy location (n = 25) measurements of PPFD were taken on each sampling date in and near the forested plot . In addition, we explored the effect of four photointensity-photoperiod combinations on the growth of L. benzoin under controlled conditions to compare to field conditions. On average, understory PPFD was less than 2% of open PPFD during the leafed months and an average of 38.8% of open PPFD during leafless winter months, indicating that: (1) often overlooked woody surfaces intercept large amounts of light; and (2) spicebush within the plot receive limited light even in early spring before canopy leaf-out. Statistical results suggested phenoseason accounted for nearly three-quarters of the variation in incident radiation between the three plant canopy heights. Spicebush under controlled conditions exhibited the highest fitness levels at an intensity of 164.5 μmol m-2 s-1 for 12-hour duration. Similarly, spicebush growth in the field occurred at subcanopy locations receiving higher incidence of PPFD . Results suggest that the ecological niche for these plants is very specific in terms of light intensity.The purpose of this work was to quantify the variation of subcanopy spatiotemporal light dynamics over the course of a year and to link it to the physiological ecology of the understory shrub,  It haThe dynamics of understory light are greatly impacted as a forest undergoes the phenological changes associated with these phenoseasons. Understory light availability, including photosynthetically active radiation PAR, 400–700 nm), has been shown to vary at all spatial and temporal scales ,6–10. Pr00–700 nm2 m-2) commonly ranging from 4 to 6 m2 m-2 during the foliated phenoseasons, light exposure is critical to understory plants which need to maximize carbon gain before the canopy leaves emerge as well as during short-lived sunflecks [It is well known that solar radiation, and PAR, decrease exponentially as a function of LAI . In deciunflecks . Althougunflecks . They tyunflecks .Lindera benzoin L. Blume (northern spicebush), is common and ecologically significant in the understory of mid-Atlantic deciduous forests. Lindera benzoin is the dominant understory species within our plot and is often found under L. tulipifera. The spicebush’s drupes are an important food source for Passerine and Phasianidae, especially during fall migration. Leaves are a food source for white-tailed deer and other herbivorous mammals, and an invaluable habitat for small to medium sized mammals, songbirds, pheasants, and insects—specifically, the namesake spicebush swallowtail and spicebush silkmoth. Many physiological traits of L. benzoin, including growth, herbivory defense, and reproduction, have been shown to be greatly affected by light intensities [L. benzoin plants of all ages is greater in sun patches and along forest edges [L. benzoin demonstrated greater amounts of new stem growth, lower stomatal density, higher photosynthetic rate, and lower leaf thickness at light levels between 191 and 391 μmol m-2 s-1 in just 11 weeks [One such plant, ensities –24. Whilensities , stem grst edges . In a st11 weeks . Leaves 11 weeks .L. benzoin, populations of spicebush swallowtail butterfly and spicebush silkmoth are both affected by the health and nutritional status of these plants, as they rely on them almost exclusively in order to survive even the first instar stage [L. benzoin, and how these cumulative differences in light exposure alter the leaf biochemistry over time, especially when healthy plants are critically needed for other native organisms such as races of papilionidae and saturniidae. How light exposure to L. benzoin changes over the course of a year is still not well understood. Answers to such questions in both the field and laboratory are necessary to a gain a holistic and more comprehensive understanding of the physiological ecology of L. benzoin in the understory.Success in the field impacts a plant’s ability to nurture insects that depend on them. In the case of ar stage . This raL. benzoin. This work is novel because it combines (1) measurements of PPFD  using a line quantum sensor in the open and at 25 locations and three elevations below the canopy across all phenoseasons with (2) a suite of laboratory experiments that examine L. benzoin response to varying photointensity- photoperiod conditions. To our knowledge, no other study has collected PAR measurements as often and at as many locations and heights, for all seven phenoseasons. This work builds upon previous studies of subcanopy radiation regimes, and contributes to gaps in the literature pertaining to PAR variability, specifically as it relates to the often under-utilized phenoseason construct, the effect of PAR availability on the physiological ecology of an ecologically significant understory plant, and measurement of subcanopy light dynamics at the microscale [in-situ, subcanopy datasets to future analyses cannot be underestimated.The aims of this study were to quantify PAR in the subcanopy and its spatial and temporal variation within and among phenoseasons in a mid-Atlantic deciduous forest, as well as to gain a better understanding of how this PAR variability affects woody understory shrubs, specifically croscale . Additio2 research plot in Fair Hill Natural Resource Management Area (NRMA) in Cecil County, MD, USA (Fagus grandifolia Ehrh. (American beech) and Liriodendron tulipifera L. (yellow poplar), with few Acer rubrum L. (red maple), Carya spp. (hickory), and Quercus spp. (oak). Average tree age within the stand is approximately 55 years. The plot contains 42 trees having a diameter at breast height (DBH) greater than 10 cm, and has a stand density of 233 trees/ha. The stand mean DBH is 39.5 ± 28.8 cm. Based on a plot inventory conducted in October of 2014, canopy heights range from 5.5 m to 30 m, with a mean height of 23 m. The stand basal area is 43.7 m2 ha-1. The plot has a woody area index of 1.21 ± 0.01 m2 m-2 and a leaf area index of 5.12 ± 0.17 m2 m-2. Soils in the area are primarily loam and silt loams of igneous, metamorphic, phyllite and schist parent material [Field work was conducted in an expanded 1800 mmaterial . Within material  with a p-2 s-1, along the length of a one-meter quartz rod conductor under an acrylic diffuser . The one-meter, fifteen-second average integration of the LI-191 has a few advantages over spot meters for this study, including size , lack of need for cosine correction (because the sensor is located under an acrylic diffuser), and most importantly, each measurement captures a much more representative sample of the overall field of PPFD .PPFD was measured using a LI-COR LI-191 Line Quantum Sensor and LI-COR LI-250A light meter , which can take an instantaneous measurement, or of recording a fifteen-second spatially-integrated average of the PPFD in units of μmol mn = 4,592). We set up the field site to collect measurements in the most timely manner to reduce changes in light conditions during collection. Most locations within the plot have a litter layer with no understory vegetation except for short-lived, seasonal flowers in the spring, and few locations have growth of ferns or jack-in-the-pulpit. Thus, it was easy to discern by sight areas in the understory where spicebush were growing densely. To better understand light availability at these locations, we framed our methodology in a way that would allow us to measure the light above, within, and under the spicebush canopy. At the five subcanopy locations within the plot where there was dense growth of L. benzoin (L. benzoin canopy (at a height of 1.75 m), within the L. benzoin canopy (0.75 m), and beneath the L. benzoin canopy (0 m) on each visit.Subcanopy PPFD measurements were acquired non-simultaneously at a height of 1.75 m, using an instrument-attached circular spirit level, at twenty-five marked locations within the plot , point 0The week, year, month, day, Julian date, phenoseason, visual sky condition, and PPFD measurements and associated times were recorded using a field sheet created this study, before being entered into a database. Visual sky conditions for each field visit were described in Oktas, a meteorological description of cloud cover in 1/8th increments of sky, where 0 is clear sky, 4 is 50% sky cloudiness, and 8 is completely overcast. To capture the broadest range of light and cloud conditions possible, trips to the field site were tracked using a spreadsheet in which the number of visits during a 15-minute interval for each phenoseason could be recorded. In this way, measurements could be collected during most daylight hours regardless of sky condition, except for instances of active precipitation. For ease of understanding, the number of collection days completed during a phenoseason’s daylight hours has been condensed into a one-hour interval in During statistical analysis of the field measurements, no outlier values were removed from the dataset because we are confident that outlier values here represent important, and often the most interesting aspects of the data. In this setting, such values represent sunflecks and spatiotemporal hotspots which can occur when radiation penetrates persistent canopy gaps for a longer duration than more highly variable sunflecks.L. benzoin seeds purchased from Horizon Herbs  were germinated in divided containers in PRO-MIX®  and grown in natural light at room temperature in the laboratory for approximately 60 days. After this time, plants were re-potted in fresh PRO-MIX® in individual pots and placed into the base of a Conviron PGC20 Reach-In Plant Growth Chamber  programmed at a constant 24°C and 68% relative humidity for 35 days for both experiments. At the time of use, the growth chamber was equipped with 400W metal halide light bulbs .To quantify spicebush adaptation to simulated light regime conditions rather than seasonality, a growth chamber was arranged for two simultaneous treatments in each of two temporally separate experiments. The first experiment was intended to simulate an exaggerated version of full-sun PAR conditions, while the second was meant to simulate more realistic subcanopy or shade conditions. Both experiments were conducted using the same set-up and temperature and humidity settings, so that the light conditions would be the only variable. For each experiment respectively, -2s-1. This value was selected based on our experience with maximum PPFD values in the subcanopy at our site, as well as a reasonable mid-point based on field work of Niesenbaum and Kluger [-2s-1 and 658.3 μmol m-2s-1 respectively. With the growth chamber programmed to 350 μmol m-2s-1 overall intensity, PPFD was manually measured at the soil surface to be 517.9 μmol m-2s-1 for the elevated upper treatment and 366 μmol m-2s-1 for the lower treatment using an LI-190 Quantum Sensor in conjunction with a LI-COR LI-250A light meter . The program was set to run 16 hours of light and 8 hours of darkness for Experiment 1. Overall photointensity for the second experiment was set to 150 μmol m-2s-1 and was intended to simulate a lower intensity, shorter photoperiod (subcanopy or shade) environment. At this intensity, the upper and lower treatments of Experiment 2 were recorded at the soil surface to be 164.5 μmol m-2s-1 and 118.06 μmol m-2s-1, respectively, for 12 hours of light and 12 hours of darkness. These values were much closer to light conditions in our field site, where we knew that average PPFD at the center of the plot just above spicebush growth was 125.79 μmol m-2s-1. The growth chamber was unable to be set for a lower photointensity than 150 μmol m-2s-1, thus limiting the lower range of PPFD for this aspect of the study.To maximize resources, each of the two treatments in the experiments were set up to receive a different photointensity for a standard photoperiod by elevating one treatment 55 cm above the other, thereby increasing the PPFD for the elevated plants. For Experiment 1, intended to simulate high-intensity and long photoperiod, the overall photointensity within the growth chamber was set to 350 μmol md Kluger who foundUsing a Model SC-1 Leaf Porometer , stomatal conductance measurements were taken on all intact leaves twice for each experiment; once during the second-to-last week in the growth chamber, and again one week later, during the last week in the growth chamber. Living growth chamber plants were non-destructively measured for leaf area, number of nodes, inter-nodal distances, and total node height. At the end of the 35-day growing stage, plants were harvested for destructive measurements including weight and dry weight of roots and shoots.t-tests among the central tendencies and dispersion of the variables. Analysis of Variance (ANOVA) was used to evaluate PAR variation in the shrub canopy. A Tukey-Kramer Honest Significant Difference test was employed to ascertain differences between plants in the growth chamber experiments. The Tukey-Kramer Honest Significant Difference test (or Tukey's Range test) is designed to identify means which differ significantly by comparing all pairs of means and utilizing a studentized  distribution [We employed statistical methods in STATISTICA 13.2 to determine differences in the PAR variations. These methods include comparing descriptive statistics and ribution . It assu-2 s-1).We compared measurements of PPFD from each location, from three canopy heights, and across seven phenoseasons. Vertical referential photographs in The spring leafless and early spring leafing phenoseasons had lower open PPFD values and higher, more variable subcanopy PPFD values than the summer phenoseasons. Late summer leafing and fully-leafed phenoseasons had PPFD values that were higher in the open, lower and less variable values in the subcanopy, with distinct hot-spots (or sunflecks) because of canopy closure, sky conditions, and the position of the sun.-2 s-1). The spring leafing and summer leafing phenoseasons had the highest standard deviations, 259.48 and 218.76 μmol m-2 s-1, respectively.Understory PPFD had, overall, a greater range of non-outlier values during the unfoliated phenoseasons, but “extreme-outlier” cases (hot spots and sunflecks) increased as median PPFD values and interquartile variability decreased with foliated phenoseasons. The summer fully-leafed and autumnal fully-leafed phenoseasons had the greatest instances of extreme values . This result was significant at the α = .05 level. When locations were ranked by mean annual PPFD, the five locations with abundant growth of L. benzoin measured in the 64th percentile of locations receiving the most solar radiation within the plot, i.e. locations receiving greater than 128 μmol m-2 s-1. When ranked by median values, four of the five locations fall within the 75th percentile. Similarly, these locations also received greater all-season cumulative flux.At the five locations within the site where Visually, the plants from Experiment 2 performed better and achieved greater fitness than the plants from Experiment 1. Mean total weight of plants from Experiment 1 was nearly half that of Experiment 2 . Plants p = 0.0130). Total dry weight of both 1U (p = 0.0007) and 1L (p = 0.0082) treatments were found to be significantly different from 2U. Plants from 2U grew to significantly higher biomass than either treatment from Experiment 1. Finally, significant differences in initial stomatal conductance were found between 1U and 1L and 2U and 2L. . In other words, there was a significant difference between the stomatal conductivity of plants grown in the 350 μmol m-2 s-1 light environment and the plants grown in the 150 μmol m-2 s-1 light environment.A Tukey-Kramer Honest Significant Difference test of non-collinear variables found the means of the root-shoot ratios and initial stomatal conductance to be significantly different (α = 0.05) for upper (1U) and lower (1L) plants of Experiment 1. Means of initial stomatal conductance for the upper (2U) and lower (2L) plants of Experiment 2 were significantly different. There was a significant difference between the root-to-shoot ratios for plants grown from 1U (highest) treatment set and plants from 2L (lowest) treatment set . Although this value is lower than what might be considered peak efficiency for this type of plant, it was a closer-to-natural scenario  than the other photoperiod-photointensity combinations. Though the lower set from Experiment 1 experienced photointensity closer to the value of that found to be conducive for peak efficiency (366 μmol m-2 s-1), understory plants do not likely experience such an intense photon flux for more than a few hours, and especially not for 16 hours straight, every day for weeks. As Chazdon and Pearcy [Complementing the field measurements, the purpose of the growth chamber treatments was to determine how changes in available light energy in the growth environment affect overall plant health. What we have shown is that in the short-term these shade-adapted plants significantly modify their physical regulation for survival in high light environments. We have also shown that over time, al genes  and the al genes ,35. Luke [et al.  showed ti.e., the combination of canopy state and celestial geometry) may account for nearly 75% of the variation between canopy heights in this mid-Atlantic deciduous forest. The influence of phenoseason on the differences of PAR received at three heights within the subcanopy, and the critical nature of its combined inputs to subcanopy plants highlights the importance of this construct to study the physiological ecology of understory plants. The other 26% may be variation caused by sky conditions, slope, aspect, and canopy species distribution. While understory PPFD is typically between 20 and 40% of open PPFD, values of PPFD in the shrub canopy are often only 5–12% of the incident PPFD. Additionally, our results agree with Hutchison and Matt’s [L. benzoin in the subcanopy by restricting a primary growth-limiting resource. Although no direct conclusions could be made about species dependencies (i.e. disparities in subcanopy light availability under L. tulipifera vs. F. grandifolia), the fact that all five subcanopy locations with greater occurrence of L. benzoin receive in the upper 36% of annual PPFD suggests that variance in the crown shape and LAI of canopy species above does impact the growth of shade-adapted species in the understory.We demonstrated that phenoseason (L. benzoin within our plot, can be to light regime changes as seemingly small as 50 μmol m-2 s-1. On a large scale, the photosynthesis of these canopy and subcanopy plants plays a large role in the carbon and water cycles, the maintenance of which are of global importance [Ecological climatology involves the bidirectional coupling of ecosystems and climate, highlighting the interconnections among populations at all scales, from individual, to stand, to regional, to global . In thesportance ,36,37. I


In this study we report antibacterial modification of Kirschner wires (K-wires) with polyluteolin (PL) toward methicillin-resistant Staphylococcus aureus (MRSA). K-wires were modified by immersing them in the luteolin-containing aqueous solution for 24 h. Characterizations using scanning electron microscopy and electrochemical methods confirmed the presence of the PL coatings on the K-wires. The PL-coated K-wires were further found to show antibacterial activity toward MRSA and remained unimpaired antibacterial activity even after the steam sterilization treatment. Orthopaedic devices used in joint and hip replacement surgeries have greatly improved the health outcomes for patients. However, implant associated infections remain a major problem in hospital settings ,2. TreatFlavonoids belong to an important group of naturally occurring phenol derivatives. Flavonoids from fruits, vegetables and cereals, herbs and spices have shown beneficial effects on human health, and have been found to be effective antimicrobial substances against a wide variety of microorganisms ,10,11. CPL-coated K-wires could be prepared by the simple immersion of the K-wires in a freshly prepared aqueous luteolin solution for 24 h, meanwhile the color of luteolin solution changed from the initial colorless to the light yellow, indicative of the spontaneous polymerization of luteolin by molecular oxygen. PL-coated K-wires were firstly examined by SEM. As revealed by vs. Ag/AgCl, indicating the formation of the PL coatings. The observed oxidation current could result from the oxidation of the catechol group on the ring B of luteolin structure and the irreversible oxidation of the resorcinol group on the ring A of luteolin structure [Then, cyclic voltammetry was further used to check the presence of PL coatings on the K-wires due to the redox properties associated with the PL coatings. tructure . During tructure ,20. ThesThe antibacterial activity of the PL-coated K-wires against MRSA was evaluated by ASTM E2149-01, a quantitative antibacterial test method for determining the antibacterial activity of the attached antibacterial agents under dynamic contact conditions. As shown in It is well known that all materials implanted within the body or placed in contact with corporeal fluids must be sterilized and steam sterilization is the preferred sterilization process . HoweverK-wires  were immersed in a freshly prepared Tris buffer  solution containing 0.10 mM luteolin  for 24 h at 37 °C.The morphology of the PL-coated K-wires was characterized using a ZEISS Supra 55 FE-SEM. Cyclic voltammetry was used to check the presence of PL coatings on the K-wires. Cyclic voltammetric measurements were performed with a CHI 660D electrochemical analyzer  with a three-electrode configuration. K-wires were used as the working electrode, Ag/AgCl (KCl-saturated) electrode as the reference electrode, and platinum coil as the counter electrode.The antimicrobial properties of the PL-coated K-wires were evaluated by ASTM E2149-01, which is a quantitative antimicrobial test method performed under dynamic contact conditions. Each specimen was sterilized before the experiment with high-pressure steam sterilizer  at 121 °C for 30 min. Gram-positive bacteria, MRSA ATCC43300, was used as test organism. Strains were stored frozen in glycerol broth at −70 °C and subcultured to ensure purity before testing.5 colony forming units (CFU) per mL) and incubated with the PL-coated K-wires at 225 rpm on a shaker for 1 and 24 h. At the end of the incubation, 100 μL solution was took out and plated onto 10-cm LB culture medium plates overnight, respectively. After 24 h incubation, the number of colonies on each plate was quantitated following protocols set forth by the U.S. Food and Drug Administration in their Bacteriological Analytical Manual-Aerobic Plate Count Method [A colony of MRSA grown on a LB agar plate was cultivated in sterilized LB broth and then incubated overnight at 37 °C with a shaking incubator. Then, bacteria were suspended in 1.5 mL culture broth (10t Method . BrieflyFor testing the effect of steam sterilization on the antimicrobial activity of the PL coatings, the PL-coated K-wires were treated with high-pressure steam sterilizer at 121 °C for 30 min, and then were incubated with 1.5 mL of cell suspension in a 2 mL conical tube at 37 °C with a shaking incubator. This steam sterilization and the subsequent incubation processes were repeated twice. Samples were taken out after 24 h, and plated on LB agar plates. The inoculated plates were incubated at 37 °C for 24 h, and the surviving cells were counted. All determinations were made in triplicate. The antimicrobial activity was expressed as R = % reduction of the organism after contact with the test samples compared to the number of S. aureus cells surviving after contact with the control.In the present study, the K-wires have been modified with the PL for producing antibacterial activity toward MRSA. Modification of the K-wires with PL coatings was facile just by immersing them in the luteolin-containing aqueous solution for 24 h. Characterization with SEM and cyclic voltammetry indicated the presence of the PL coatings on the K-wires. The PL-coated K-wires were found to possess good antibacterial activity toward MRSA. Moreover, such antibacterial activity was unimpaired by the steam sterilization, which will facilitate the practical application of the PL-coated K-wires.


Psychotria henryi. Accompanying with significant activation of cyclic electron flow (CEF), the P700 oxidation ratio was maintained at high levels when exposed to strong light either under fluctuating light or constant high light. Under moderate fluctuating light, PSI and PSII activities were remained stable in P. henryi. Interestingly, PSI was insusceptible to fluctuating light but sensitive to constant high light in P. henryi. Furthermore, both PSI and PSII were more sensitive to constant high light than fluctuating light. These results suggest that CEF is essential for photoprotection of PSI under fluctuating light in P. henryi. Furthermore, photoinhibition of PSI under high light in P. henryi is more related to the accumulation of reactive oxygen species rather than to P700 redox state, which is different from the mechanisms of PSI photoinhibition in Arabidopsis thaliana and rice. Taking together, PSI is a key determiner of photosynthetic responses to fluctuating light and constant high light in the shade-establishing species P. henryi.Shade-establishing plants growing in the forest understory are exposed to constant high light or fluctuating light when gaps are created by fallen trees. Our previous studies indicate that photosystem I (PSI) is sensitive to constant high light in shade-establishing tree species, however, the effects of fluctuating light on PSI and photosystem II (PSII) in shade-establishing species are little known. In the present study, we examined the responses of PSI and PSII to fluctuating light in comparison to constant high light in the shade-establishing species  Psychotria rubra . When canopy gaps are created by fallen trees, leaves of shade-establishing plants may be exposed to fluctuating light or constant high light for several hours in a day. Under constant high light, excess light energy usually causes selective photodamage to photosystem II (PSII) . Generalia rubra , 2016d. Arabidopsis thaliana   , 2004. Iactivity . Previough light , 2004. RI to PSI , which nox state . These rA. thaliana  was chosen for study. P. henryi is a shade-establishing shrub species of rain forests native to south of China, and the maximum height of its plants is about 2 m. In our present study, plants of P. henryi grown naturally in forest understory with deep shade (light intensity < 5% of sunlight) in tropical rain forest in Xishuangbanna tropical botanical garden  were used for photosynthetic measurements. After cutting off from plants, the branches were immediately immersed in water and brought back to our lab in 5 min, and the photosynthetic measurements were conducted subsequently.The tropical tree species Fv/Fm = (Fm–Fo)/Fm, was measured to monitor PSII activity. Other fluorescence parameters were calculated as follows (Y(II) = (s)/m–Fs)/(Fs). Fm is the minimum and maximum fluorescence measured after 30 min dark adaptation. Fs is the steady-state fluorescence after light adaptation. Fm and -2 s-1). Y(II) represents the effective quantum yield of PSII, NPQ indicates the non-photochemical quenching. qL indicates the fraction of PSII centers in the open state (with oxidized plastoquinone).In our present study, the PSI and PSII parameters were measured simultaneously by Dual-PAM-100 . After dark adaptation for 30 min, the minimum and maximum fluorescence and the maximum photo-oxidizable P700 were measured. The maximum quantum yield of PSII,  follows : Y(II) =+ signals (P) may vary between minimal (P700 is fully reduced) and maximal levels (P700 is fully oxidized). The maximum level (Pm) was determined with application of a saturation pulse (300 ms and 10000 μmol photons m-2 s-1) after pre-illumination with far-red light  = (P)/Pm, and the P700 oxidation ratio in a given actinic light was calculated as Y(ND) = P/Pm. The fraction of P700 that cannot be oxidized by SP to the overall P700 was calculated as Y(NA) =  . The P70ed light , 2008. Pble P700 . In our Y(I), and ETRII = PPFD × 0.5 × abs I ×Y(II), where 0.5 is the fraction of absorbed light reaching PSI or PSII, and abs I is absorbed irradiance taken as 0.84 of incident irradiance.The electron transport rate (ETR) was calculated as ETRI = PPFD × 0.5 × abs I ×-2 s-1 for 2 min and high light at 1294 μmol photons m-2 s-1 for 2 min) for 60 min. PSI and PSII parameters were measured every 1 min during this period. Before and after photoinhibitory treatments, the PSI activity (Pm) and the PSII activity (Fv/Fm) were measured after 30 min dark adaptation.In the present study, light from a 635 nm light-emitting diode (LED) equipped in Dual-PAM-100 was used as actinic light for photoinhibitory treatments. In order to examine the response of PSI and PSII to moderate fluctuating light, intact leaves were dark-adapted for 30 min and then were exposed to fluctuating light  for 100 min. Afterward, leaves were dark-adapted for 30 min and the values of Pm and Fv/Fm were measured. Immediately, these leaves were exposed to the above fluctuating light or constant high light for 100 min, and then Pm and Fv/Fm were measured after 30 min dark adaptation.To further determine the effects of intense fluctuating light on PSI and PSII activities, values for The results were displayed as mean values of five independent experiments. One-Way ANOVA test was used at α = 0.05 significance level to determine whether significant differences existed between different treatments.-2 s-1 every 2 min for 60 min, values of Y(I), Y(II), ETRI, ETRII were constant at 100 μmol photons m-2 s-1 and gradually increased in the initial 12 min when exposed to 1294 μmol photons m-2 s-1 . NPQ quickly increased from the low levels at 100 μmol photons m-2 s-1 to the high levels at 1294 μmol photons m-2 s-1 in 1 min , suggesting the rapid generation of ΔpH. The value of 1 – qL was maintained at high levels at high-light phases and subsequently decreased to low levels at low-light phases , indicating the rapid regulation of the redox state of the plastoquinone pool. Similarly, the P700 redox state under fluctuating light was regulated quickly in response to changes in light conditions. Y(ND) fast increased to high levels during the high-light phases and decreased to low levels during the subsequent low-light phases . Similarly, Y(NA) rapidly decreased to low levels during the high-light phases .Under fluctuating light alternating between 100 and 1294 μmol photons m-2 s-1. However, ETRI was two times higher than ETRII when illuminated at 1294 μmol photons m-2 s-1 . These results suggested that the shade-establishing species P. henryi showed significantly activation of CEF around PSI under high light, which was confirmed by the high levels of Y(ND) and low levels of Y(NA) at high light. Furthermore, the rapid changes in NPQ, 1 – qL, Y(ND), and Y(NA) further implied the physiological roles of CEF under fluctuating light. Interestingly, after exposure to this moderate fluctuating light (100 μmol photons m-2 s-1 for 2 min and 1294 μmol photons m-2 s-1 for 2 min) for 60 min, both Pm and Fv/Fm were remained stable , indicating that PSI and PSII activities were insusceptible to short time moderate fluctuating light in the shade-establishing tree species P. henryi.The value of ETRI approximately equaled that of ETRII at the low light of 100 μmol photons m-2 s-1 compared with the treatment with constant light of 2258 μmol photons m-2 s-1 . Because NPQ is calculated as NPQ = . However, this increase in NPQ is not truth owing to the decrease in Fm after high light treatment. After treatment with fluctuating light or constant for 100 min, the actual value of NPQ at 2258 μmol photons m-2 s-1 largely decreased, as shown by comparison between NPQ values at 5 and 135 min . Under fluctuating light, NPQ can be activated quickly during the high-light phases, suggesting the activation of CEF. During the treatment period of 0–100 min at fluctuating light, the value of Y(II) at 94 μmol photons m-2 s-1 changed little . However, during the treatment period of 100–200 min, the value of Y(II) at low-light phases gradually decreased with increasing time . The redox state of the plastoquinone pool (1 – qL) changed slightly during the treatments with constant high light , suggesting the accumulation of reducing power in photosynthetic electron transport system. By comparison, the redox state of the plastoquinone pool was rapidly regulated under fluctuating light, indicating that the accumulated reducing power during the high-light phases can be dissipated during the subsequent low-light phases.The value of Y(I) during treatments with fluctuating light and constant light was similar to Y(II) . Y(ND) changed regularly according to light condition under fluctuating light and gradually decreased under constant high light treatment . The value of Y(NA) at 2258 μmol photons m-2 s-1 increased gradient with increasing time of treatments under both fluctuating and constant light . Notably, the increase in Y(NA) under constant high light was faster than under fluctuating light. Similar to Y(ND), Y(NA) was regulated regularly under fluctuating light.The trend of change in -2 s-1 every 5 min for 100 and 200 min, the value of Pm only decreased by 4 and 7%, respectively . Meanwhile, the value of Fv/Fm decreased to 0.62 and 0.49 . By comparison, after treatment with constant light (2258 μmol photons m-2 s-1) for 100 and 200 min, Pm decreased by 11 and 24%, respectively . Concomitantly, Fv/Fm decreased to 0.40 and 0.24, respectively . These results strongly indicate that (1) PSI is insusceptible to fluctuating light in P. henryi; (2) both PSI and PSII are more sensitive to constant high light than fluctuating light in P. henryi; (3) photoinhibition of PSI under constant high light cannot be prevented by the activation of CEF.After exposure to fluctuating light alternating between 2258 and 94 μmol photons m2 assimilation rate, plant biomass . However, constant high light led to significant decrease in PSI activity in P. henryi . These results strongly indicate that PSI is a key determiner of photosynthetic acclimation to high light under fluctuating light or constant high light in the shade-establishing species P. henryi. Furthermore, photoinhibition of PSI and PSII was significantly stronger after treatment with constant high light than fluctuating light in P. henryi , which is different from the phenomenon in model plant A. thaliana . Photoinhibition of PSII occurs only when the rate of photodamage exceeds the rate of repair. After exposure to moderate fluctuating light (100 and 1294 μmol photons m-2 s-1 every 2 min) for 60 min, Fv/Fm only showed a little decrease in P. henryi , indicating that the photodamage induced by high-light (1294 μmol photons m-2 s-1) phases can be fast recovered at the subsequent low-light (100 μmol photons m-2 s-1) phases. Furthermore, this result suggested that exposure to 1294 μmol photons m-2 s-1 for 2 min hardly caused significant accumulation of ROS, because the repair of photodamaged PSII can be inhibited by ROS , indicating that the low-light phases significantly affect the extent of PSII photoinhibition. As indicated in Figure 1, the maximum value of ETRII was approximately 30 μmol electrons m-2 s-1, indicating the relatively low ability to utilize light energy in P. henryi. Furthermore, the value of 1 – qL was maintained at high levels when exposed to high light , suggesting that the plastoquinone pool was over-reduced due to accumulation of reducing power in the photosynthetic electron transport system. As a result, PSII in P. henryi was very sensitive to constant high light , owing to excess production of ROS . As a result, the low-light phases favor the scavenging of ROS and the fast repair of PSII. Additionally, the rate of PSII photodamage is dependent on light intensity , and Y(NA) . However, it also showed significantly PSI photoinhibition after treatment with constant high light . Furthermore, recent studies found that PSI photoinhibition also occurred in wild-type of A. thaliana  is essential for photosynthesis in angiosperm plants . As indident CEF . Under cn of ΔpH , which fn of ΔpH , 2016a,bgh light , but alsgh light . Therefogh light . On the thaliana  and ricethaliana  after exP. rubra . These rP. henryi , whereas short-term fluctuating light hardly inactivated PSI activity . As a result, PSI is a key determiner of photosynthetic responses to fluctuating light and constant high light in P. henryi. These results are largely opposite to the phenomenon in the pgr5 plants of both A. thaliana and rice, in which PSI is more susceptible to fluctuating light than to constant high light . As a result, unlike pgr5 plants of A. thaliana, PSI photoinhibition is less related to P700 oxidation ratio in P. henryi. This result is consistent with our recent studies on a coexisting relative P. rubra species, in which PSI photoinhibition under high light is mainly determined by the electron flow from PSII to PSI , indicating that the reducing power that accumulates during high-light phases can be dissipated quickly during the subsequent low-light phases. As a result, the ROS production was diminished during the low-light phases, and then over-accumulation of ROS was avoided in P. henryi, which was confirmed by the stability of PSI and PSII after treatment with moderate fluctuating light for 60 min . Under constant high light, the total incident light energy that leaves receive is much higher than that receive under fluctuating light. And the over-reduction of photosynthetic electron chains, as indicated by high levels of 1 – qL, induced the production and accumulation of ROS and then aggravated PSII photoinhibition . Based on the results of stronger PSI and PSII photoinhibition under constant high light than under fluctuating light , we propose that PSI photoinhibition in P. henryi is largely dependent on the accumulation of ROS. In other words, the two coexisting relative shade-establishing species P. henryi and P. rubra probably have the same mechanism of PSI photoinhibition under high light.Under high light, P700 redox state plays an important role in determining the sensitivity of PSI to photoinhibition in model plant thaliana . In pgr5gh light . In the I to PSI . Under fP. henryi. The significant PSI photoinhibition under high light indicated that ROS generated under high light could not be scavenged quickly by anti-oxidative enzymes, leading to accumulation of ROS. This implies the insufficient capacity of the anti-oxidative system in the chloroplast in shade-establishing plants. As a result, future studies are still needed to examine the relationship between high-light-induced PSI photoinhibition and the capacity of anti-oxidative system in shade-establishing species. Although PSI photoinhibition is inevitable in the shade-establishing species P. henryi and P. rubra, PSI photoinhibition also represent some kind of protective mechanisms against long lasting over-reduction of PSI acceptor side, diminishing production of enormous quantity of ROS and preventing extensive damage of cell structures and cell death in stress conditions  across the thylakoid membranes within 1 min , prevents PSI and PSII photoinhibition from moderate fluctuating light (100 and 1294 μmol photons m-2 s-1 every 2 min for 60 min). These results suggest that CEF is critical for photosynthetic acclimation to fluctuating light in both light-demanding and shade-establishing plants. Under constant high light, PSI photoinhibition can be avoided owing to the activation of CEF in light-demanding plants. However, CEF cannot completely prevent PSI photoinhibition in shade-establishing species such as P. henryi  and P. rubra  strongly indicated that shade-establishing species have the ability to avoid photoinhibition during the period of short-term light spot. The significantly stronger photoinhibition of PSI and PSII under constant high light indicated that the mechanisms of PSI photoinhibition in P. henryi are different from A. thaliana and rice. In P. henryi, PSI photoinhibition is more related to the accumulation of ROS rather than to P700 redox state, which is similar to its relative P. rubra species (In the present study, we examined the responses of PSI and PSII to fluctuating light and constant high light in the shade-establishing tropical tree species  species . FurtherWH and S-BZ conceived and designed research. WH and Y-JY conducted experiments. WH and Y-JY analyzed data. WH, HH, and S-BZ wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.The reviewer MR and handling Editor declared their shared affiliation, and the handling Editor states that the process nevertheless met the standards of a fair and objective review.


This meta-analysis was conducted to compare the intraoperative and postoperative outcomes of bimanual microincision cataract surgery (B-MICS) and coaxial microincision cataract surgery (C-MICS).  Three databases were searched for papers that compared B-MICS and C-MICS from inception to June 2016. The following intraoperative and postoperative outcomes were included in the final meta-analysis: ultrasound time (UST), effective phacoemulsification time (EPT), balanced salt solution use (BSS use), mean surgery time, best-corrected visual acuity (BCVA), central corneal thickness (CCT), and increased CCT. p < 0.01) and lower levels of increased CCT at postoperative day 1 (p = 0.02) in the B-MICS group compared with the C-MICS group.  There were no statistically significant differences in mean surgery time, UST, BSS use, BCVA, CCT, or increased CCT (one subgroup at postoperative day 7-8 and another subgroup at postoperative day 30). However, there was less EPT needed during surgery ( The EPT was shorter and increased CCT was less at postoperative day 1 in the B-MICS group. There were no statistically significant differences in other intraoperative and postoperative outcomes between the B-MICS group and the C-MICS group. B-MICS is an efficient and safe cataract surgery procedure. With the development of equipment for cataract surgery and increased requirements for visual outcome, the main recent change in cataract surgical procedure aims to decrease the size of the clean corneal incision. In coaxial microincision cataract surgery (C-MICS), irrigation, aspiration, and phacoemulsification are performed with the same instruments, which is similar to standard coaxial small incision cataract surgery (C-SICS) but provides a smaller incision , 2. LessAccording to a recent study , there aThis meta-analysis was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement .The following terms were searched in PubMed, Web of Science, and the Cochrane Library: “bimanual microincision,” “biaxial microincision,” “bimanual microincisional cataract surgery,” “bimanual phacoemulsification,” “biaxial phacoemulsification,” “coaxial microincision,” “coaxial microincisional cataract surgery,” and “coaxial phacoemulsification.” Papers published before July 2016 were included in the search. The references of the included papers were reviewed to seek papers that were missed in the primary search. The papers included in the meta-analysis met the following criteria: (1) the incision of the coaxial microincision cataract surgery (C-MICS) was less than 2.2 mm and (2) patients in the study had no other ocular diseases other than cataracts. Two investigators (Fu and Chu) searched the three databases in all fields independently.After scanning the titles and abstracts, Fu and Chu obtained access to the full text of the papers that compared outcomes of C-MICS and B-MICS. After reading the full text, papers were included in the meta-analysis based on the criteria listed above. The search and exclusion process was conducted as shown in p < 0.01. For heterogeneity among studies, the significance level was set at p < 0.10 for Cochran's Q statistic and I2 > 50% for the I2 index score [2 score > 50%), in which case, a random-effects model based on the DerSimonian and Laird method was used [The data on all outcomes were analysed using Stata software . The means and SDs of outcomes were extracted from the papers to obtain the weighted mean difference (WMD) with a 95% confidence interval (CI). Except for heterogeneity and metaregression analyses of the data, the statistical significance level was set at was used . To assewas used . The staAmong the included papers, there were seven randomized controlled trials (RCTs) , 14–19 ap ≤ 0.001, I2 = 49.3%, Pheterogeneity = 0.066).In seven studies reporting on 589 eyes, the EPT was longer in the C-MICS group than in the B-MICS group in the fixed-effects model .In four studies that reported on 330 eyes, the mean surgery time was recorded. The forest plot showed that there was no statistically significant difference between the B-MICS group and the C-MICS group in the random-effects model .As shown in p = 0.177, I2 = 83.6%, Pheterogeneity = 0.002).Three studies reported UST, and significant heterogeneity in UST among these studies was found. As shown in p = 0.735, I2 = 0.0%, Pheterogeneity = 0.570; at day 30: WMD: −0.003, 95% CI: −0.021 to 0.014, p = 0.697, I2 = 8.9%, Pheterogeneity = 0.334).In five studies, the BCVA was measured at postoperative day 1, day 7, day 30, day 60, and day 90. Subgroup meta-analysis was performed for two subgroups: BCVA within 7 postoperative days (222 eyes) and BCVA at day 30 (160 eyes). In both groups, no statistically significant difference was found between the B-MICS group and the C-MICS group in the fixed-effects model .CCT was measured in three studies. We used data from two studies that recorded the precise CCT measuring time for 82 and 90 eyes, respectively, and analysed the postoperative CCT in the day 1 subgroup and the after day 30 subgroup. As shown in the forest plot, no statistically significant difference was found in either subgroup .In two studies, one reporting on 60 eyes and one reporting on 90 eyes, increased CCT was measured and calculated. Three subgroups  were assessed. There were no statistically significant differences in the day 7-8 subgroup or in the day 30 subgroup. And for the subgroup day 1, the forest plot showed that increased CCT was less common in the B-MICS group than in the C-MICS group in the fixed-effects model .No publication biases were found for any of the results of the intraoperative and postoperative outcomes.For the pooled results of the B-MICS and C-MICS in this meta-analysis, no statistically significant differences were found in mean surgery time, UST, BSS use, postoperative BCVA (within 7 days and at day 30), postoperative CCT (at day 1 and after day 30), and postoperative increased CCT (at day 7-8 and day 30). The shorter EPT in B-MICS may be due to the “cold” phacoemulsification mode used in B-MICS, or it may be due to the separation of the irrigation port from the aspiration port in B-MICS, which can avoid a competing current from the phacoemulsification tip and assist in the emulsification and aspiration process. Moreover, the separation and exchangeable hand pieces can provide surgeons with more flexibility to clean the subincisional cortex and residual viscoelastic material , 19, 22.The intraoperative and postoperative complications were not analysed because the incidence of intraoperative complications was quite low, and long-term postoperative complications such as posterior capsule opacification (PCO) were rarely found during follow-up in the included studies. PCO develops several months to a few years after uneventful cataract surgeries, but the longest follow-up time in the included studies was three months. Among all cases reported in the studies, three cases in the B-MICS group (6.6%) and one (2.2%) in the C-MICS group from the Can et al. study suffered from intraoperative complications . In all Four papers included in this meta-analysis reported SIA , 16, 18,Because the incision sizes of B-MICS and C-MICS are both micro, SIA or BCVA may not be precise enough to detect differences between the two procedures. In three studies , 17, 18,Although the incision of cornea is 2.0 mm or less in the original definition of MICS , 31, a fThere are some advantages to our study. All nine studies included in this meta-analysis were prospective, and seven of the studies were randomized trials. The operations in the B-MICS and C-MICS groups were performed by the same experienced surgeon in the majority of the included studies , 14–18, The learning curve is also considered to be a drawback of the pervasiveness of B-MICS. Some related studies have been conducted. In one long-term follow-up study comparing the clinical results, such as PCO incidence and clear corneal incision (CCI) architecture of B-MICS, between surgeons in training and experienced surgeons , the PCOIn this meta-analysis, there was a shorter EPT and lower levels of increased CCT at postoperative day 1 in the B-MICS group, but other main clinical outcomes of B-MICS and C-MICS were not significantly different. Thus, both cataract surgery techniques are efficient, safe, and appropriate for cataract surgery.


EditorialMYC gene resulting in its overexpression and cell transformation [MYC gene to an immunoglobulin locus, which in B cells would result in constitutive expression [MYC was long considered a diagnostic criterion for BL, recent evidence suggests other mechanisms may be responsible for MYC deregulation in a small percentage of BL cases [Burkitt lymphoma (BL) is a B-cell malignancy which occurs with varying clinical manifestations and frequencies. The most well-known presentation of BL is found in areas which are holoendemic or hyperendemic for malaria. This endemic BL occurs primarily in children and presents as tumors of the jaw ,2. Thougormation ,8. Most BL cases .+ T-cell activation is weak and lasting cellular immunity is not generated. The HLA class II response remains an area of interest, however, as antigen presentation through class II molecules, resulting in activation of CD4+ T-cells, has been shown to enhance and sustain cytotoxic T lymphocyte (CTL) immunity [BL is a rapidly growing malignancy with a doubling time of 24 hours and consequently requires aggressive therapies for treatment . While cimmunity . While timmunity ,14.+ T-cell activation: BL lines Nalm-6, Ramos and Ous, and B-LCL lines Frev and 6.16.DR4.DM. Cell lines were retrovirally transduced to generate: Ramos.DR4, Nalm-6.DR4, and 6.16.DR4.DM; Ous and Frev constitutively express HLA-DR4 molecules. Surface expression of HLA-DR4 was confirmed in all cell lines by flow cytometry [+ T-cells through the HLA-DR4 class II molecule while 6.16.DR4.DM generated a strong CD4+ T-cell response [vs pH 5.5). When antigen presentation assays with 6.16.DR4.DM were performed in the eluate obtained from incubating BL cells in pH 5.5 buffer, it was discovered that the >30 kDa fraction of the pH 5.5 eluate drastically impaired the antigen presentation ability of 6.16.DR4.DM. Further, this impairment was ablated by treating the eluate with proteinase K. Taken together, this suggests the presence of Burkitt Lymphoma-Associated Inhibitory Molecule (BLAIM) which is proteinaceous in nature.To explore this, we employed several strategies in the hopes of isolating inhibitory molecules which may be implicated in the HLA class II antigen presentation defect. We have employed several cell lines for the purpose of assaying CD4ytometry . Thus, aresponse ,14. PeptIn an effort to shed light on the identity of BLAIM, we next compared protein expression patterns between Frev, Nalm-6.DR4 and 6.16.DR4.DM cells. pH 5.5 eluates obtained from each cell line were separated on a non-reducing gel and the gel was then stained with Coomassie blue. A comparison of the banding pattern revealed a ~50 kDa protein which was highly expressed in Frev and 6.16.DR4.Dm but absent or only slightly expressed in Nalm-6.DR4. Extraction and mass spectrophotometric analysis of this protein band revealed a 47 kDa enolase-like protein . Furtherc-MYC is overexpressed display a reduced capacity to present antigen via HLA class II molecules [high and c-MYClow cells. Our results demonstrated that while surface expression of HLA class II is not decreased in c-MYChigh cells when compared to c-MYClow cells, the intracellular expression of two critical components of the class II pathway were sharply decreased. We observed a significant decrease in the expression of HLA-DM and GILT  in c-MYChigh cells when compared to c-MYClow cells. HLA-DM and GILT are both essential for the efficient and proper processing of antigens for presentation via the HLA class II pathway. It logically follows that decreasing the expression levels of either one of these molecules would have deleterious effect on the ability of cells to present antigens via class II molecules [In support of our idea that the defect in HLA class II antigen presentation is multifactorial, we have previously shown that cells in which olecules . To inveolecules .c-MYC overexpression and decreased expression of class II pathway processing components. A greater understanding of either the presentation or processing defects could lead to the development of novel immunotherapies which could be used in support of traditional chemotherapy, or could eliminate the need for chemotherapy altogether. The development of alternative and less toxic therapies is a necessary endeavor for those patient populations who do not tolerate well the aggressive treatments traditionally used for BL. Further, given the similarities among many cancers, we are hopeful that our work with BL will provide insights which may lead to a greater understanding of the immunological aspects of cancer development.Thus, we have demonstrated that BL cells appear to have multiple points of defect in their ability to effectively present antigen through the HLA class II pathway. The process of antigen presentation is hampered both at the level of antigen processing, resulting from decreased levels of class II pathway antigen processing components, and at the level of antigen presentation, through BLAIM and the 47 kDa enolase-like protein which appear to interfere directly with presentation. Ongoing and future studies focus on a more detailed characterization of BLAIM and the 47 kDa enolase-like protein. Additionally, we will continue to explore the link between